bloodthirsty, an RBCC/TRIM gene required for erythropoiesis in zebrafish.
The Antarctic icefishes (family Channichthyidae, suborder Notothenioidei) constitute the only vertebrate taxon that fails to produce red blood cells. These fishes can be paired with closely related, but erythrocyte-producing, notothenioids to discover erythropoietic genes via representational difference analysis. Using a B30.2-domain-encoding DNA probe so derived from the hematopoietic kidney (pronephros) of a red-blooded Antarctic rockcod, Notothenia coriiceps, we discovered a related, novel gene, bloodthirsty (bty), that encoded a 547-residue protein that contains sequential RING finger, B Box, coiled-coil, and B30.2 domains. bty mRNA was expressed by the pronephric kidney of N. coriiceps at a steady-state level 10-fold greater than that found in the kidney of the icefish Chaenocephalus aceratus. To test the function of bty, we cloned the orthologous zebrafish gene from a kidney cDNA library. Whole-mount in situ hybridization of zebrafish embryos showed that bty mRNA was present throughout development and, after the mid-blastula transition, was expressed in the head and in or near the site of primitive erythropoiesis in the tail just prior to red cell production. One- to four-cell embryos injected with two distinct antisense morpholino oligonucleotides (MOs) targeted to the 5'-end of the bty mRNA failed to develop red cells, whereas embryos injected with 4- and 5-bp mismatch control MOs produced wild-type quantities of erythrocytes. The morphant phenotype was rescued by co-injection of synthetic bty mRNA containing an artificial 5'-untranslated region (UTR) with the antisense MO that bound the 5'-UTR of the wild-type bty transcript. Furthermore, the expression of genes that mark terminal erythroid differentiation was greatly reduced in the antisense-MO-treated embryos. We conclude that bty is likely to play a role in differentiation of the committed red cell progenitor.
Yergeau
Donald A
DA
Department of Biology, Northeastern University, 360 Huntington Avenue, Boston, MA 02115, USA.
Cornell
Catharine N
CN
Parker
Sandra K
SK
Zhou
Yi
Y
Detrich
H William
HW
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics*; D002454:Cell Differentiation; D017931:DNA Primers; D018076:DNA, Complementary / Q000235:genetics; D004625:Embryo, Nonmammalian / Q000502:physiology*; D004920:Erythropoiesis / Q000502:physiology*; D018507:Gene Expression Regulation, Developmental; D017403:In Situ Hybridization; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity; D014158:Transcription, Genetic; D015027:Zebrafish / Q000196:embryology*; D029961:Zebrafish Proteins / Q000737:chemistry / Q000235:genetics*
D002352:Carrier Proteins; D017931:DNA Primers; D018076:DNA, Complementary; D029961:Zebrafish Proteins; C503418:bty protein, zebrafish