Two-domain hemoglobin gene of the water flea Moina macrocopa: duplication in the ancestral Cladocera, diversification, and loss of a bridge intron.
Two cDNAs encoding the two-domain hemoglobin (Hb) chains of a crustacean Cladocera, Moina macrocopa, were cloned and their nucleotide (nt) sequences were determined. The amino acid (aa) sequences of both the gene products deduced from the nt sequences consisted of 348 residues and showed 98% identity with each other. These sequences together with the NH(2)-terminal aa sequences of the Hb chains determined after separation by two-dimensional gel electrophoresis showed that the Hb chains are synthesized as a secretory precursor with a signal peptide of 17 aa residues. The aa sequences of M. macrocopa Hb chains shared the following features with those of Daphnia Hb chains. Firstly, the signal peptide is followed by an NH(2)-terminal extension containing a threonine-rich sequence that might play a role in the multimerization of subunit chains. Secondly, the identity between the aa sequences of the first and second domains is exceptionally low. These facts suggest that duplication of the cladoceran Hb gene occurred before the divergence of families Moinidae and Daphniidae. Analysis of genomic DNA showed that the M. macrocopa Hb genes consist of two large repeated regions, encoding the first and second domains of Hb chains, respectively. The intron-exon organization of the first region of the M. macrocopa Hb genes was similar to that found in the Daphnia Hb genes, having the three-exon, two-intron structure characteristic of animal Hb genes. However, the intron bridging the two regions and the most downstream intron in the second region were missing in the Moina genes, providing a new example of intron loss. The following elements in the 5' flanking region were conserved in the Moina and Daphnia genes: (1) TATAAA, a typical TATA box sequence accompanied by a downstream sequence, GAAXAGCATCAGTT (the fourth residue X was G or A in Daphnia and absent in Moina); (2) CCAAT boxes, located upstream of the TATA box; (3) the binding sites for HIF-1 and GATA-1, also located upstream of the TATA box, that may be responsible for up-regulation of the cladoceran Hb genes under hypoxia.
Kato
K
K
Laboratory of Environmental and Molecular Biology, Environmental Science Division, School of Life Science, Tokyo University of Pharmacy and Life Science, 1432-1 Horinouchi, Hachioji, Tokyo 192-0392, Japan.
Tokishita
S
S
Mandokoro
Y
Y
Kimura
S
S
Ohta
T
T
Kobayashi
M
M
Yamagata
H
H
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D017124:Conserved Sequence; D003445:Crustacea / Q000235:genetics*; D015180:Electrophoresis, Gel, Two-Dimensional; D019143:Evolution, Molecular; D020440:Gene Duplication; D006454:Hemoglobins / Q000235:genetics*; D007438:Introns; D010802:Phylogeny; D017434:Protein Structure, Tertiary; D012045:Regulatory Sequences, Nucleic Acid; D017385:Sequence Homology
D006454:Hemoglobins