Conservation of a master hematopoietic switch gene during vertebrate evolution: isolation and characterization of Ikaros from teleost and amphibian species.
The generation of T, B and NK lymphocyte lineages from pluripotent hematopoietic stem cells is dependent upon the early expression of the Ikaros locus which by means of alternative splicing produces a variety of zinc finger DNA binding transcription factors. We assessed the general biological importance of Ikaros by studying its conservation and expression in teleost fish and amphibians. Portions of Ikaros cDNA from rainbow trout and Xenopus were amplified by reverse transcription-polymerase chain reaction (RT-PCR). They show roughly 75% conservation of the amino acid sequence with mammalian Ikaros. The trout fragment was then used to isolate full-length Ikaros clones from a trout thymocyte cDNA library. In mice and humans, Ikaros produces six alternatively spliced isoforms, but in trout two additional novel splice variants designated Ik-7 and Ik-8 were also found. Ik-7 is expressed in a similar fashion to Ik-1 and Ik-2, the predominant isoforms expressed in mammalian lymphocytes. In trout and zebrafish, as in mammals, Ikaros is a single-copy gene, but in Xenopus segregation analysis demonstrates that Ikaros has been duplicated, most likely a result of polyploidization. We then examined the expression of Ikaros in trout and Xenopus tumor T cell lines via Northern blot, RT-PCR, immunofluorescence and Western blot analysis. Overall, Ikaros is expressed in a lymphoid-specific fashion similar to that found in mice and humans. In addition Ikaros is expressed early in trout ontogeny, beginning roughly at days 3-4 in the yolk sac and at day 5-6 in the embryo proper. The conservation of Ikaros structure and expression confirms it as a master switch of hematopoiesis.
Hansen
J D
JD
Basel Institute for Immunology, Switzerland. hansen@bii.ch
Strassburger
P
P
Du Pasquier
L
L
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D017124:Conserved Sequence; D018076:DNA, Complementary / Q000302:isolation & purification; D004268:DNA-Binding Proteins; D019143:Evolution, Molecular; D006412:Hematopoietic Stem Cells / Q000276:immunology* / Q000378:metabolism*; D006801:Humans; D051740:Ikaros Transcription Factor; D017578:Immunoglobulin Class Switching / Q000235:genetics*; D008221:Lymphoid Tissue / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D017686:Oncorhynchus mykiss; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000096:biosynthesis; D017386:Sequence Homology, Amino Acid; D014157:Transcription Factors / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification*; D014981:Xenopus; D029961:Zebrafish Proteins; D016335:Zinc Fingers / Q000235:genetics
D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D012333:RNA, Messenger; D014157:Transcription Factors; D029961:Zebrafish Proteins; C497238:Zfpn1a1 protein, mouse; C497240:ikzf1 protein, zebrafish; D051740:Ikaros Transcription Factor