title: The RHOX homeobox gene cluster is selectively expressed in human oocytes and male germ cells.
abstract: OBJECTIVE
  What human tissues and cell types express the X-linked reproductive homeobox (RHOX) gene cluster?
  
  
  CONCLUSIONS
  The RHOX homeobox genes and proteins are selectively expressed in germ cells in both the ovary and testis.
  
  
  BACKGROUND
  The RHOX homeobox transcription factors are encoded by an X-linked gene cluster whose members are selectively expressed in the male and female reproductive tract of mice and rats. The Rhox genes have undergone strong selection pressure to rapidly evolve, making it uncertain whether they maintain their reproductive tissue-centric expression pattern in humans, an issue we address in this report.
  
  
  METHODS
  We examined the expression of all members of the human RHOX gene cluster in 11 fetal and 8 adult tissues. The focus of our analysis was on fetal testes, where we evaluated 16 different samples from 8 to 20 weeks gestation. We also analyzed fixed sections from fetal testes, adult testes and adult ovaries to determine the cell type-specific expression pattern of the proteins encoded by RHOX genes.
  
  
  METHODS
  We used quantitative reverse transcription-polymerase chain reaction analysis to assay human RHOX gene expression. We generated antisera against RHOX proteins and used them for western blotting, immunohistochemical and immunofluorescence analyses of RHOXF1 and RHOXF2/2B protein expression.
  
  
  RESULTS
  We found that the RHOXF1 and RHOXF2/2B genes are highly expressed in the testis and exhibit low or undetectable expression in most other organs. Using RHOXF1- and RHOXF2/2B-specific antiserum, we found that both RHOXF1 and RHOXF2/2B are primarily expressed in germ cells in the adult testis. Early stage germ cells (spermatogonia and early spermatocytes) express RHOXF2/2B, while later stage germ cells (pachytene spermatocytes and round spermatids) express RHOXF1. Both RHOXF1 and RHOXF2/2B are expressed in prespermatogonia in human fetal testes. Consistent with this, RHOXF1 and RHOXF2/2B mRNA expression increases in the second trimester during fetal testes development when gonocytes differentiate into prespermatogonia. In the human adult ovary, we found that RHOXF1 and RHOXF2/2B are primarily expressed in oocytes.
  
  
  CONCLUSIONS
  While the average level of expression of RHOX genes was low or undetectable in all 19 human tissues other than testes, it is still possible that RHOX genes are highly expressed in a small subset of cells in some of these non-testicular tissues. As a case in point, we found that RHOX proteins are highly expressed in oocytes within the human ovary, despite low levels of RHOX mRNA in the whole ovary.
  
  
  CONCLUSIONS
  The cell type-specific and developmentally regulated expression pattern of the RHOX transcription factors suggests that they perform regulatory functions during human fetal germ cell development, spermatogenesis and oogenesis. Our results also raise the possibility that modulation of RHOX gene levels could correct some cases of human infertility and that their encoded proteins are candidate targets for contraceptive drug design.
authors.affiliation: Department of Reproductive Medicine, School of Medicine, University of California San Diego, 9500 Gilman Drive, La Jolla, CA 92093, USA.
authors.forename: H W
authors.initials: HW
authors.lastname: Song
authors.forename: R A
authors.initials: RA
authors.lastname: Anderson
authors.forename: R A
authors.initials: RA
authors.lastname: Bayne
authors.forename: J
authors.initials: J
authors.lastname: Gromoll
authors.forename: S
authors.initials: S
authors.lastname: Shimasaki
authors.forename: R J
authors.initials: RJ
authors.lastname: Chang
authors.forename: M M
authors.initials: MM
authors.lastname: Parast
authors.forename: L C
authors.initials: LC
authors.lastname: Laurent
authors.forename: D G
authors.initials: DG
authors.lastname: de Rooij
authors.forename: T C
authors.initials: TC
authors.lastname: Hsieh
authors.forename: M F
authors.initials: MF
authors.lastname: Wilkinson
mesh_terms: D000328:Adult; D000595:Amino Acid Sequence; D015153:Blotting, Western; D005260:Female; D018507:Gene Expression Regulation, Developmental; D018398:Homeodomain Proteins / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D008297:Male; D008875:Middle Aged; D008969:Molecular Sequence Data; D005810:Multigene Family; D009865:Oocytes / Q000378:metabolism*; D010920:Placenta / Q000378:metabolism; D011247:Pregnancy; D011261:Pregnancy Trimester, First; D011262:Pregnancy Trimester, Second; D013094:Spermatozoa / Q000378:metabolism*; D013737:Testis / Q000378:metabolism
chemical_list: D018398:Homeodomain Proteins; C585950:RHOXF1 protein, human; C471968:RHOXF2 protein, human
keywords: homeobox gene cluster; oogenesis; ovary; spermatogenesis; testis