Expression of the Ladybird-like homeobox 2 transcription factor in the developing mouse testis and epididymis
© Moisan et al; licensee BioMed Central Ltd. 2008
Received: 15 July 2007
Accepted: 27 February 2008
Published: 27 February 2008
Homeoproteins are a class of transcription factors that are well-known regulators of organogenesis and cell differentiation in numerous tissues, including the male reproductive system. Indeed, a handful of homeoproteins have so far been identified in the testis and epididymis where a few were shown to play important developmental roles. Through a degenerate PCR approach aimed at identifying novel homeoproteins expressed in the male reproductive system, we have detected several homeoproteins most of which had never been described before in this tissue. One of these homeoproteins is Ladybird-like homeobox 2 (Lbx2), a homeobox factor mostly known to be expressed in the nervous system.
To better define the expression profile of Lbx2 in the male reproductive system, we have performed in situ hybridization throughout testicular and epididymal development and into adulthood. Lbx2 expression was also confirmed by real time RT-PCR in those tissues and in several testicular and epididymal cell lines. In the epididymis, a highly segmented tissue, Lbx2 shows a regionalized expression profile, being more expressed in proximal segments of the caput epididymis than any other segment. In the testis, we found that Lbx2 is constitutively expressed at high levels in Sertoli cells. In interstitial cells, Lbx2 is weakly expressed during fetal and early postnatal life, highly expressed around P32-P36, and absent in adult animals. Finally, Lbx2 can also be detected in a population of germ cells in adults.
Altogether, our data suggest that the homeoprotein Lbx2 might be involved in the regulation of male reproductive system development and cell differentiation as well as in male epididymal segmentation.
Homeobox genes encode transcription factors known as homeoproteins that share a highly conserved 60 amino acid DNA-binding motif called a homeodomain [1–3]. Homeoproteins are known to regulate expression of genes involved in critical developmental and physiological processes in all living organisms. These processes include body plan segmentation, organogenesis, molecular gradient specification, and cell lineage specification and differentiation. Homeoproteins have been identified in several tissues and the male reproductive system is no exception (reviewed in [4, 5]).
The male reproductive system is essential for the production of fully functional gametes and for the establishment of the secondary sexual characteristics. It is composed of the testis and several secondary sex organs: the rete testis, epididymis, vas deferens, seminal vesicles, prostate and bulbourethral glands. Proper development of the male reproductive system is thus indispensable for normal male sex differentiation and reproductive function. The process of male sex determination/differentiation is triggered by the Y chromosome-linked SRY (Sex-determining Region Y) gene (reviewed in ). In the mouse, Sry is transiently expressed (between embryonic day 10.5 and E12.5) specifically in pre-Sertoli cells. Since SRY expression is limited to a discrete period of testis differentiation [7, 8], it acts as a molecular switch to turn on a network of molecular and cellular events essentials for testicular development as well as male sex differentiation. Three critical hormones produced by the somatic cells of the newly formed testis are essential for male sex differentiation and reproductive function: Müllerian inhibiting substance/anti-Müllerian hormone (MIS/AMH), insulin-like 3 (INSL3), and testosterone (reviewed in ). MIS, a hormone belonging to the TGFβ family, is produced by Sertoli cells and regulates male sex differentiation by triggering regression of the Müllerian ducts, which if left intact would develop into the internal female reproductive tract (fallopian tubes, uterus, and upper part of the vagina) . Testosterone, a steroid hormone, secreted by Leydig cells and its more potent derivative dihydrotestosterone regulate several key processes that include testicular descent, development of the accessory sex glands and external genitalia, masculinization of the brain, male sexual behavior, and initiation and maintenance of spermatogenesis (male gamete production) . INSL3, a small peptide belonging to the insulin/relaxin/growth factor family also produced by Leydig cells, regulates the first phase of testis descent during fetal life [11, 12] and acts as a germ cell survival factor in adults .
Although the testis is the site of spermatogenesis, spermatozoa that exit the testis do not have the capacity to fertilize eggs. The final steps of spermatozoa maturation (acquisition of motility, chromatin condensation) occur in the epididymis, a convoluted and androgen-regulated organ composed of one long tubule divided into three distinct regions called caput, corpus and cauda. Epididymal segmentation is directly related to its function which is species-conserved [14, 15]. In addition to functionality, each region of the epididymal tubule is characterized by a distinct physiology. Therefore several genes have been shown to have a region-specific expression along the epididymis tubule . In addition to its importance for sperm maturation, the epididymis also serves as a reservoir for spermatozoa .
The process of testis and epididymis formation, as for organogenesis of all tissues, relies on a network of hormones and signaling molecules that act by regulating expression of genes involved in specifying the unique features and functions of these tissues. Some of these genes encode transcription factors. In recent years, some homeoproteins have been implicated in testicular and epididymal development and include Emx2 , Lhx9 , Pbx1 , Arx , HoxA10 , and Pax2 .
Here we report the identification, through a degenerate PCR approach, of additional homeobox factors expressed in the male reproductive system. In addition, we have performed a detailed characterization of the expression profile during testicular and epididymal development of one of the homeoprotein identified, the Ladybird-like homeobox 2 (Lbx2) homeoprotein.
Results and Discussion
Homeoproteins identified by degenerate PCR in Leydig cells and epididymis
Hoxb-2, Hoxb-3, Hoxb-9
Lbx2 is the second member of a family that also comprises Lbx1 and Lbx3. The mammalian Lbx1 and Lbx2 gene are the homologs of the Drosophila Ladybird genes Ladybird late (Lbl) and Ladybird early (Lbe). Ladybird-like genes were also identified in the chick embryo, Lbx1 and Lbx3, which share a high degree of homology with mammalian Lbx genes . In Drosophila, Lbl and Lbe have been shown to play important roles in neurogenesis, myogenesis, and cardiogenesis [30–33]. A consensus DNA binding site for Lbl and Lbe proteins, RVYTAAYHAG, was recently identified . This motif was then used in a ChIP-enriched in silico target approach (ChEST) that led to the identification of several target genes regulated by the Ladybird factors in Drosophila . These genes were found to encode proteins involved in cardiac and muscle cell fate specification as well as in cell shape, adhesion, and motility . Interestingly, in mammals Lbx1 was reported to play equally important roles. Indeed, Lbx1-/- mice have important defects in heart looping , interneuron specification in the spinal cord [36–38], and migration of muscle cell precursors [39–41]. As for Lbx3, its role in chick remains unknown and no mammalian homolog has been identified yet.
Although Lbx2 was found to be strongly expressed in both tissues throughout development into adulthood, we did not detect expression of the highly related Lbx family member Lbx1 (data not shown). This is consistent with the fact that Lbx1 and Lbx2 do not have overlapping expression patterns in general [24, 54] and are therefore believed to play non-redundant roles during development. The expression pattern of Lbx2 in the testis and epididymis described herein supports the notion that this transcription factor might be involved in the development and/or the function of these organs. While this manuscript was in preparation, Lbx2 null mice have been reported . Surprisingly, Lbx2-/- mice are viable and show no gross morphological defects and both male and female Lbx2-/- mice were found to be fertile, although no detailed analyses of the reproductive system were reported . A mild partial lethality associated with Lbx2 deficient mice was however observed but failed to reach statistical significance . As suggested by Wei et al, back crossing the Lbx2 mutation in a different genetic background may be required to detect a phenotype associated with Lbx2 deficiency . Besides the genetic background, this lack of a penetrant phenotype might be explained by a redundancy mechanism where another homeobox factor could compensate for the absence of Lbx2. This is very common amongst genes that are essential for development and cell differentiation, including homeobox encoding genes [55–57]. Wei et al proposed that the Tlx2 homeobox gene is an attracting candidate as a substitute for Lbx2 for several reasons. First, Tlx2 and Lbx2 belong to the superclass of homeobox proteins . Second, the genomic location and organization of the Tlx2 and Lbx2 genes are conserved in Drosophila and mice . And finally, Tlx2 and Lbx2 expression patterns are overlapping in numerous tissues including the testis [24, 26, 59]. Besides Tlx2, it is also possible that other yet unidentified Lbx family members could also compensate for the absence of Lbx2. In agreement with this is the identification of Lbx3 in the avian genome, although a mammalian homolog has yet to be identified . If it does indeed exist, this other Lbx family member would represent an interesting candidate to compensate for the absence of Lbx2.
In conclusion, our present study provides new insights into the expression profile of the homeobox factor Lbx2 throughout development of the testis and the epididymis. The lack of overt phenotype in Lbx2 null mice may indicate that Lbx2 does not play a dominant role in the development and the function of these organs. Another possibility is that other homeobox factors compensate for the absence of Lbx2. Since Lbx2 expression is dynamic in the testis and epididymis, Lbx2 constitutes a useful molecular marker for histological and developmental studies.
C57BL/6 mice were maintained on a 12L:12D light cycle with water and food ad libitum. Mice were killed at different time points as indicated in the figure legends and the testes and epididymides were harvested. Whole testis and epididymis were fixed in 4% (w/v) paraformaldehyde for 24 h. Tissues were then dehydrate with ethanol, substituted with xylene, and embedded in paraffin. All experiments complied with the regulations set by the Animal Welfare Act (Public Law 91-579), the Canadian Council for Animal Care, the Guide for the Care and Use of Laboratory Animals (National Research Council, 1996) published by the Department of Health and Human Services, and the policies and procedures of the University of Virginia Institutional Animal Care and Use Committee. All experiments have been approved by the Animal Care and Ethics Committee of Laval University (protocol # 06-059).
Most cell lines used in the present study were obtained from ATCC (Leydig: mTLC-1, TM3, R2C; and Sertoli: TM4, 15P-1). The MA-10 Leydig cell line  was provided by Dr. Mario Ascoli (University of Iowa, Iowa City, IA) and the Sertoli MSC-1 cell line was a gift from Dr. Michael Griswold (Washington State University, Pullman, WA). The MSC-1 cell lines were grown in Dulbecco modified Eagle medium (DMEM) supplemented with 10% fetal bovine serum, HEPES and 50 mg/liter of penicillin and streptomycin sulfates. MA-10, were grown in Waymouth's MB752/1 medium supplemented with 20 mM HEPES, 15% horse serum and 50 mg/liter of penicillin and streptomycin sulfates. All cell lines obtained from ATCC were cultured as recommended by ATCC. Cell lines were grown at 37°C and 5% CO2.
RNA preparation and RT-PCR
Sequence-specific primers used in the RT-PCR studies
Three different probes for Lbx2 were tested. A 604 bp fragment that encompasses the entire coding sequence (nt 60–624 of Genbank accession number NM_010692), a 504 bp fragment from nt 120 to 624, and a 263 bp fragment that contains the coding sequence C-terminal of the homeodomain into the 3' UTR (nt 490–753). The fragments were obtained by PCR and cloned into pBluescript (Stratagene). Sense and antisense digoxigenin (DIG)-labeled riboprobes for Lbx2 were subsequently obtained by linearizing the plasmid followed by in vitro transcription using T7 or T3 RNA polymerase (GE Healthcare) in the presence of DIG-UTP (Roche Diagnostics, Laval, Canada). The DIG-labeled riboprobes were then used in in situ hybridization experiments on paraformaldehyde-fixed, paraffin-embedded tissue sections. In brief, testis and epididymis sections were dewaxed in xylene, rehydrated in graded alcohols (95%, 70%, and 50%) and diethylpyrocarbonate-treated water, and digested by proteinase K (10 mg/mL) for 15 min. Glycine (2 mg/mL) was used to stop the proteinase K digestion. Tissues were then refixed with 4% paraformaldehyde and treated with 0.25% acetic anhydride in 0.1% triethanolamine (pH 8.0) for 10 min. Between each step, the slides were washed twice in PBS (pH 7.5) for 5 min. The sections were then prehybridized in hybridization solution (0.3 M NaCl; 10 mM Tris-HCl, pH7.5; 1 mM EDTA; 1× Denhardt's; 5% dextran sulfate; 0.02% sodium dodecyl sulfate; 50% formamide; and 250 μg/ml salmon sperm DNA) at 42°C for 16 hrs and finally hybridized in 30 μl of the same solution containing 7.5 μg/mL DIG-labeled Lbx2 antisense or sense riboprobe at 42°C. On the next day, the slides were washed twice for 10 min at 42°C with 2 × SSC, 1 × SSC, 0.2 × SSC and 0.05 × SSC and incubated with a 1:1000 dilution of an alkaline phosphatase-conjugated anti-DIG antiserum (Roche Diagnostics, Laval, Canada) for 2 h at room temperature. Nitroblue tetrazolium chloride and 5-bromo-4-chloro-3-indolylphosphate p-toluidine (NBT/BCIP) were used as substrates for the alkaline phosphatase reaction. Sections were counterstained with 5% neutral red and mounted in Permount (Fisher Scientific, Montreal, Canada). The results presented were obtained with the 504 bp probe.
Paraformaldehyde-fixed, paraffin-embedded testis sections were dewaxed in xylene, treated 30 min in 0.3% H2O2 (Sigma-Aldrich, Oakville, Canada)/methanol, rehydrated in graded alcohols (95%, 70%, and 50%) and treated for antigen retrieval. Sections were then blocked for 2 h with 10% horse serum and incubated overnight at 4°C with an Anti-Müllerian inhibitory substance antiserum (MIS, 1:100, Santa Cruz Biotechnology) in PBS containing 0.1% BSA. The next morning, the slides were washed in PBS and incubated 45 min with a biotinylated anti-goat antibody (1:1500, Vector Laboratories, Burlington, Canada). After washing in PBS, sections were submitted to an avidin-biotin complex (ABC) solution for 20 min at room temperature (Vectastain ABC Elite Kit, Vector Laboratories, Burlington, Canada). The signal was detected using a solution of 3-amino-9-ethylcarbazole (AEC, Sigma-Aldrich Canada, Oakville, Canada), 50 mM acetate buffer pH 5.2 (0.2 M sodium acetate; 0.2 M acetic acid) and 0.002% H2O2. Sections were then counterstained with Gill #1 hematoxylin and mounted in 15% glycerol and 0.1% sodium azide in PBS.
We would like to thank Dr. Matthew Hardy (Population Council, NY) for generously providing RNA from purified Leydig cells. We are indebted to Drs. Mario Ascoli, and Michael Griswold, for providing the MA-10 and MSC-1 cell lines, respectively. JJT holds a New Investigator scholarship from the Canadian Institutes of Health Research (CIHR). This work was supported by CIHR grant number MOP-81387 to JJT.
- Desplan C, Theis J, O'Farrell PH: The Drosophila developmental gene, engrailed, encodes a sequence-specific DNA binding activity. Nature. 1985, 318: 630-635. 10.1038/318630a0.View ArticlePubMed CentralPubMedGoogle Scholar
- Desplan C, Theis J, O'Farrell PH: The sequence specificity of homeodomain-DNA interaction. Cell. 1988, 54: 1081-1090. 10.1016/0092-8674(88)90123-7.View ArticlePubMed CentralPubMedGoogle Scholar
- Hoey T, Levine M: Divergent homeo box proteins recognize similar DNA sequences in Drosophila. Nature. 1988, 332: 858-861. 10.1038/332858a0.View ArticlePubMedGoogle Scholar
- Bomgardner D, Hinton BT, Turner TT: Hox transcription factors may play a role in regulating segmental function of the adult epididymis. J Androl. 2001, 22: 527-531.PubMedGoogle Scholar
- Lindsey JS, Wilkinson MF: Pem: a testosterone- and LH-regulated homeobox gene expressed in mouse Sertoli cells and epididymis. Dev Biol. 1996, 179: 471-484. 10.1006/dbio.1996.0276.View ArticlePubMedGoogle Scholar
- Viger RS, Silversides DW, Tremblay JJ: New insights into the regulation of mammalian sex determination and male sex differentiation. Vitam Horm. 2005, 70: 387-413. 10.1016/S0083-6729(05)70013-3.View ArticlePubMedGoogle Scholar
- Hacker A, Capel B, Goodfellow P, Lovell-Badge R: Expression of Sry, the mouse sex determining gene. Development. 1995, 121: 1603-1614.PubMedGoogle Scholar
- Koopman P, Münsterberg A, Capel B, Vivian N, Lovell-Badge R: Expression of a candidate sex-determining gene during mouse testis differentiation. Nature. 1990, 348: 450-452. 10.1038/348450a0.View ArticlePubMedGoogle Scholar
- Teixeira J, Maheswaran S, Donahoe PK: Mullerian inhibiting substance: an instructive developmental hormone with diagnostic and possible therapeutic applications. Endocr Rev. 2001, 22: 657-674. 10.1210/er.22.5.657.PubMedGoogle Scholar
- Roy AK, Chatterjee B: Androgen action. Crit Rev Eukaryot Gene Expr. 1995, 5: 157-176.View ArticlePubMedGoogle Scholar
- Nef S, Parada LF: Cryptorchidism in mice mutant for Insl3. Nat Genet. 1999, 22: 295-299. 10.1038/10364.View ArticlePubMedGoogle Scholar
- Zimmermann S, Steding G, Emmen JM, Brinkmann AO, Nayernia K, Holstein AF, Engel W, Adham IM: Targeted disruption of the Insl3 gene causes bilateral cryptorchidism. Mol Endocrinol. 1999, 13: 681-691. 10.1210/me.13.5.681.View ArticlePubMedGoogle Scholar
- Kawamura K, Kumagai J, Sudo S, Chun SY, Pisarska M, Morita H, Toppari J, Fu P, Wade JD, Bathgate RA, Hsueh AJ: Paracrine regulation of mammalian oocyte maturation and male germ cell survival. Proc Natl Acad Sci U S A. 2004, 101: 7323-7328. 10.1073/pnas.0307061101.View ArticlePubMed CentralPubMedGoogle Scholar
- Turner TT, Bomgardner D, Jacobs JP, Nguyen QA: Association of segmentation of the epididymal interstitium with segmented tubule function in rats and mice. Reproduction. 2003, 125: 871-878. 10.1530/rep.0.1250871.View ArticlePubMedGoogle Scholar
- Jones RC: Evolution of the vertebrate epididymis. J Reprod Fertil Suppl. 1998, 53: 163-181.PubMedGoogle Scholar
- Kirchhoff C: Gene expression in the epididymis. Int Rev Cytol. 1999, 188: 133-202. 10.1016/S0074-7696(08)61567-3.View ArticlePubMedGoogle Scholar
- Jones RC: To store or mature spermatozoa? The primary role of the epididymis. Int J Androl. 1999, 22: 57-67. 10.1046/j.1365-2605.1999.00151.x.View ArticlePubMedGoogle Scholar
- Miyamoto N, Yoshida M, Kuratani S, Matsuo I, Aizawa S: Defects of urogenital development in mice lacking Emx2. Development. 1997, 124: 1653-1664.PubMedGoogle Scholar
- Birk OS, Casiano DE, Wassif CA, Cogliati T, Zhao L, Zhao Y, Grinberg A, Huang S, Kreidberg JA, Parker KL, Porter FD, Westphal H: The LIM homeobox gene Lhx9 is essential for mouse gonad formation. Nature. 2000, 403: 909-913. 10.1038/35002622.View ArticlePubMedGoogle Scholar
- Schnabel CA, Selleri L, Cleary ML: Pbx1 is essential for adrenal development and urogenital differentiation. Genesis. 2003, 37: 123-130. 10.1002/gene.10235.View ArticlePubMedGoogle Scholar
- Kitamura K, Yanazawa M, Sugiyama N, Miura H, Iizuka-Kogo A, Kusaka M, Omichi K, Suzuki R, Kato-Fukui Y, Kamiirisa K, Matsuo M, Kamijo S, Kasahara M, Yoshioka H, Ogata T, Fukuda T, Kondo I, Kato M, Dobyns WB, Yokoyama M, Morohashi K: Mutation of ARX causes abnormal development of forebrain and testes in mice and X-linked lissencephaly with abnormal genitalia in humans. Nat Genet. 2002, 32: 359-369. 10.1038/ng1009.View ArticlePubMedGoogle Scholar
- Podlasek CA, Seo RM, Clemens JQ, Ma L, Maas RL, Bushman W: Hoxa-10 deficient male mice exhibit abnormal development of the accessory sex organs. Dev Dyn. 1999, 214: 1-12. 10.1002/(SICI)1097-0177(199901)214:1<1::AID-DVDY1>3.0.CO;2-2.View ArticlePubMedGoogle Scholar
- Torres M, Gomez-Pardo E, Dressler GR, Gruss P: Pax-2 controls multiple steps of urogenital development. Development. 1995, 121: 4057-4065.PubMedGoogle Scholar
- Chen F, Liu KC, Epstein JA: Lbx2, a novel murine homeobox gene related to the Drosophila ladybird genes is expressed in the developing urogenital system, eye and brain. Mech Dev. 1999, 84: 181-184. 10.1016/S0925-4773(99)00073-8.View ArticlePubMedGoogle Scholar
- Ohtoshi A, Nishijima I, Justice MJ, Behringer RR: Dmbx1, a novel evolutionarily conserved paired-like homeobox gene expressed in the brain of mouse embryos. Mech Dev. 2002, 110: 241-244. 10.1016/S0925-4773(01)00587-1.View ArticlePubMedGoogle Scholar
- Wei K, Chen J, Akrami K, Sekhon R, Chen F: Generation of mice deficient for Lbx2, a gene expressed in the urogenital system, nervous system, and Pax3 dependent tissues. Genesis. 2007, 45: 361-368. 10.1002/dvg.20302.View ArticlePubMedGoogle Scholar
- Bouma GJ, Hart GT, Washburn LL, Recknagel AK, Eicher EM: Using real time RT-PCR analysis to determine multiple gene expression patterns during XX and XY mouse fetal gonad development. Gene Expr Patterns. 2004, 5: 141-149. 10.1016/j.modgep.2004.05.001.View ArticlePubMedGoogle Scholar
- Pellegrini M, Pantano S, Lucchini F, Fumi M, Forabosco A: Emx2 developmental expression in the primordia of the reproductive and excretory systems. Anat Embryol (Berl). 1997, 196: 427-433. 10.1007/s004290050110.View ArticleGoogle Scholar
- Kanamoto T, Terada K, Yoshikawa H, Furukawa T: Cloning and expression pattern of lbx3, a novel chick homeobox gene. Gene Expr Patterns. 2006, 6: 241-246. 10.1016/j.modgep.2005.08.004.View ArticlePubMedGoogle Scholar
- Jagla K, Frasch M, Jagla T, Dretzen G, Bellard F, Bellard M: ladybird, a new component of the cardiogenic pathway in Drosophila required for diversification of heart precursors. Development. 1997, 124: 3471-3479.PubMedGoogle Scholar
- Jagla K, Jagla T, Heitzler P, Dretzen G, Bellard F, Bellard M: ladybird, a tandem of homeobox genes that maintain late wingless expression in terminal and dorsal epidermis of the Drosophila embryo. Development. 1997, 124: 91-100.PubMedGoogle Scholar
- Jagla T, Bellard F, Lutz Y, Dretzen G, Bellard M, Jagla K: ladybird determines cell fate decisions during diversification of Drosophila somatic muscles. Development. 1998, 125: 3699-3708.PubMedGoogle Scholar
- De GF, Jagla T, Daponte JP, Rickert C, Dastugue B, Urban J, Jagla K: The ladybird homeobox genes are essential for the specification of a subpopulation of neural cells. Dev Biol. 2004, 270: 122-134. 10.1016/j.ydbio.2004.02.014.View ArticleGoogle Scholar
- Junion G, Bataille L, Jagla T, Da Ponte JP, Tapin R, Jagla K: Genome-wide view of cell fate specification: ladybird acts at multiple levels during diversification of muscle and heart precursors. Genes Dev. 2007, 21: 3163-3180. 10.1101/gad.437307.View ArticlePubMed CentralPubMedGoogle Scholar
- Schafer K, Neuhaus P, Kruse J, Braun T: The homeobox gene Lbx1 specifies a subpopulation of cardiac neural crest necessary for normal heart development. Circ Res. 2003, 92: 73-80. 10.1161/01.RES.0000050587.76563.A5.View ArticlePubMedGoogle Scholar
- Muller T, Brohmann H, Pierani A, Heppenstall PA, Lewin GR, Jessell TM, Birchmeier C: The homeodomain factor lbx1 distinguishes two major programs of neuronal differentiation in the dorsal spinal cord. Neuron. 2002, 34: 551-562. 10.1016/S0896-6273(02)00689-X.View ArticlePubMedGoogle Scholar
- Kruger M, Schafer K, Braun T: The homeobox containing gene Lbx1 is required for correct dorsal-ventral patterning of the neural tube. J Neurochem. 2002, 82: 774-782. 10.1046/j.1471-4159.2002.01078.x.View ArticlePubMedGoogle Scholar
- Gross MK, Dottori M, Goulding M: Lbx1 specifies somatosensory association interneurons in the dorsal spinal cord. Neuron. 2002, 34: 535-549. 10.1016/S0896-6273(02)00690-6.View ArticlePubMedGoogle Scholar
- Brohmann H, Jagla K, Birchmeier C: The role of Lbx1 in migration of muscle precursor cells. Development. 2000, 127: 437-445.PubMedGoogle Scholar
- Gross MK, Moran-Rivard L, Velasquez T, Nakatsu MN, Jagla K, Goulding M: Lbx1 is required for muscle precursor migration along a lateral pathway into the limb. Development. 2000, 127: 413-424.PubMedGoogle Scholar
- Schafer K, Braun T: Early specification of limb muscle precursor cells by the homeobox gene Lbx1h. Nat Genet. 1999, 23: 213-216. 10.1038/13843.View ArticlePubMedGoogle Scholar
- Dyche WJ: A comparative study of the differentiation and involution of the Mullerian duct and Wolffian duct in the male and female fetal mouse. J Morphol. 1979, 162: 175-209. 10.1002/jmor.1051620203.View ArticlePubMedGoogle Scholar
- Staack A, Donjacour AA, Brody J, Cunha GR, Carroll P: Mouse urogenital development: a practical approach. Differentiation. 2003, 71: 402-413. 10.1046/j.1432-0436.2003.7107004.x.View ArticlePubMedGoogle Scholar
- Sharpe RM, McKinnell C, Kivlin C, Fisher JS: Proliferation and functional maturation of Sertoli cells, and their relevance to disorders of testis function in adulthood. Reproduction. 2003, 125: 769-784. 10.1530/rep.0.1250769.View ArticlePubMedGoogle Scholar
- O'Shaughnessy PJ, Baker PJ, Johnston H: The foetal Leydig cell - differentiation, function and regulation. Int J Androl. 2006, 29: 90-95. 10.1111/j.1365-2605.2005.00555.x.View ArticlePubMedGoogle Scholar
- Benton L, Shan LX, Hardy MP: Differentiation of adult Leydig cells. J Steroid Biochem Mol Biol. 1995, 53: 61-68. 10.1016/0960-0760(95)00022-R.View ArticlePubMedGoogle Scholar
- Mendis-Handagama SM, Ariyaratne HB: Differentiation of the adult Leydig cell population in the postnatal testis. Biol Reprod. 2001, 65: 660-671. 10.1095/biolreprod65.3.660.View ArticlePubMedGoogle Scholar
- Favier B, Dolle P: Developmental functions of mammalian Hox genes. Mol Hum Reprod. 1997, 3: 115-131. 10.1093/molehr/3.2.115.View ArticlePubMedGoogle Scholar
- Bomgardner D, Hinton BT, Turner TT: 5' hox genes and meis 1, a hox-DNA binding cofactor, are expressed in the adult mouse epididymis. Biol Reprod. 2003, 68: 644-650. 10.1095/biolreprod.102.009324.View ArticlePubMedGoogle Scholar
- Drevet JR, Lareyre JJ, Schwaab V, Vernet P, Dufaure JP: The PEA3 protein of the Ets oncogene family is a putative transcriptional modulator of the mouse epididymis-specific glutathione peroxidase gene gpx5. Mol Reprod Dev. 1998, 49: 131-140. 10.1002/(SICI)1098-2795(199802)49:2<131::AID-MRD4>3.0.CO;2-Q.View ArticlePubMedGoogle Scholar
- Maclean JA, Chen MA, Wayne CM, Bruce SR, Rao M, Meistrich ML, Macleod C, Wilkinson MF: Rhox: a new homeobox gene cluster. Cell. 2005, 120: 369-382. 10.1016/j.cell.2004.12.022.View ArticlePubMedGoogle Scholar
- Benson GV, Lim H, Paria BC, Satokata I, Dey SK, Maas RL: Mechanisms of reduced fertility in Hoxa-10 mutant mice: uterine homeosis and loss of maternal Hoxa-10 expression. Development. 1996, 122: 2687-2696.PubMedGoogle Scholar
- Hsieh-Li HM, Witte DP, Weinstein M, Branford W, Li H, Small K, Potter SS: Hoxa 11 structure, extensive antisense transcription, and function in male and female fertility. Development. 1995, 121: 1373-1385.PubMedGoogle Scholar
- Jagla K, Dolle P, Mattei MG, Jagla T, Schuhbaur B, Dretzen G, Bellard F, Bellard M: Mouse Lbx1 and human LBX1 define a novel mammalian homeobox gene family related to the Drosophila lady bird genes. Mech Dev. 1995, 53: 345-356. 10.1016/0925-4773(95)00450-5.View ArticlePubMedGoogle Scholar
- Prince VE, Pickett FB: Splitting pairs: the diverging fates of duplicated genes. Nat Rev Genet. 2002, 3: 827-837. 10.1038/nrg928.View ArticlePubMedGoogle Scholar
- Lappin TR, Grier DG, Thompson A, Halliday HL: HOX genes: seductive science, mysterious mechanisms. Ulster Med J. 2006, 75: 23-31.PubMed CentralPubMedGoogle Scholar
- Martienssen R, Irish V: Copying out our ABCs: the role of gene redundancy in interpreting genetic hierarchies. Trends Genet. 1999, 15: 435-437. 10.1016/S0168-9525(99)01833-8.View ArticlePubMedGoogle Scholar
- Holland PW: Beyond the Hox: how widespread is homeobox gene clustering?. J Anat. 2001, 199: 13-23.View ArticlePubMed CentralPubMedGoogle Scholar
- Hatano M, Iitsuka Y, Yamamoto H, Dezawa M, Yusa S, Kohno Y, Tokuhisa T: Ncx, a Hox11 related gene, is expressed in a variety of tissues derived from neural crest cells. Anat Embryol (Berl). 1997, 195: 419-425. 10.1007/s004290050061.View ArticleGoogle Scholar
- Ascoli M: Characterization of several clonal lines of cultured Leydig tumor cells: gonadotropin receptors and steroidogenic responses. Endocrinology. 1981, 108: 88-95.View ArticlePubMedGoogle Scholar
- Ge RS, Dong Q, Sottas CM, Chen H, Zirkin BR, Hardy MP: Gene Expression in Rat Leydig Cells During Development from the Progenitor to Adult Stage: A Cluster Analysis. Biol Reprod. 2005, 72: 1405-1415. 10.1095/biolreprod.104.037499.View ArticlePubMedGoogle Scholar
- Guigon CJ, Coudouel N, Mazaud-Guittot S, Forest MG, Magre S: Follicular cells acquire sertoli cell characteristics after oocyte loss. Endocrinology. 2005, 146: 2992-3004. 10.1210/en.2005-0045.View ArticlePubMedGoogle Scholar
This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.