title: A dominant-negative isoform of hypoxia-inducible factor-1 alpha specifically expressed in human testis.
abstract: Spermatogenesis in the seminiferous tubuli of the testis occurs under a high proliferation rate, suggesting considerable oxygen consumption. Because of the lack of blood vessels, the oxygen partial pressure in the lumen of these tubuli is very low. We previously identified a testis isoform of the hypoxia-inducible factor (HIF)-1alpha in the mouse, termed mHIF-1alphaI.1. Here, we demonstrate that expression of mHIF-1alphaI.1 increases during puberty, further demonstrating its gene induction in postmeiotic germ cells. Using 5'-rapid amplification of cDNA ends, we identified a novel HIF-1alpha isoform in the human testis, called hHIF-1alphaTe. Like mHIF-1alphaI.1, hHIF-1alphaTe mRNA is derived from an alternative promoter-first exon combination, but with a different genomic organization and a different nucleotide sequence. Reverse transcription-polymerase chain reaction analysis confirmed that hHIF-1alphaTe is exclusively expressed in the testis. As determined by immunofluorescence of ejaculated sperm cells, HIF-1alpha protein is mainly localized in the postacrosomal head and in the midpiece of spermatozoa. Though overlapping with mitochondrial localization in human and mouse spermatozoa, neither hHIF-1alphaTe nor hHIF-1alpha associated with mitochondria. In contrast with the ubiquitously expressed HIF-1alpha protein and the mouse testis-specific mHIF-1alphaI.1 isoform, the hHIF-1alphaTe mRNA sequence predicts a protein with an N-terminal truncation of the DNA-binding domain. As shown by yeast two-hybrid assays, hHIF-1alphaTe still formed heterodimeric complexes with HIF-1beta. However, hHIF-1alphaTe was incapable of forming a DNA-binding HIF-1 complex. Overexpression of exogenous hHIF-1alphaTe resulted in the inhibition of the endogenous HIF-1 transcriptional activity, demonstrating that the testis-specific hHIF-1alphaTe isoform is a dominant-negative regulator of normal HIF-1 activity.
authors.affiliation: Institute of Physiology, University of Lübeck, Lübeck, Germany.
authors.forename: Reinhard
authors.initials: R
authors.lastname: Depping
authors.forename: Sonja
authors.initials: S
authors.lastname: Hägele
authors.forename: Klaus F
authors.initials: KF
authors.lastname: Wagner
authors.forename: Rudolf J
authors.initials: RJ
authors.lastname: Wiesner
authors.forename: Gieri
authors.initials: G
authors.lastname: Camenisch
authors.forename: Roland H
authors.initials: RH
authors.lastname: Wenger
authors.forename: Dörthe M
authors.initials: DM
authors.lastname: Katschinski
mesh_terms: D000375:Aging / Q000378:metabolism; D000818:Animals; D001483:Base Sequence; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism* / Q000502:physiology; D005784:Gene Amplification; D005799:Genes, Dominant; D006801:Humans; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism* / Q000502:physiology; D020033:Protein Isoforms / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D013094:Spermatozoa / Q000378:metabolism; D013737:Testis / Q000254:growth & development / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism* / Q000502:physiology
chemical_list: D004268:DNA-Binding Proteins; C497442:HIF1A protein, human; C497443:Hif1a protein, mouse; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D009687:Nuclear Proteins; D020033:Protein Isoforms; D012333:RNA, Messenger; D014157:Transcription Factors; D004247:DNA