The liver of Fundulus heteroclitus expresses deiodinase type 1 mRNA.
The presence of a type 1 deiodinase (D1) in the liver of teleosts has been a controversial issue. Recently we characterized the deiodinase activity in rainbow trout and killifish liver and found that the liver of both species co-expresses the two enzymes (D1 and D2) that catalyze the outer ring-deiodinating pathway. We here report the cloning and characterization of an mRNA from the liver of the killifish Fundulus heteroclitus that encodes a D1 (FhD1). The cDNA amplified by RT-PCR from F. heteroclitus liver is 1314 nt long and encodes a protein of 248 aa. It contains a TGA codon in its open reading frame and a selenocysteine insertion sequence in its 3(') untranslated region, consistent with the structure of a selenoenzyme mRNA. The deduced peptide sequence is 73% identical to that encoded by the tilapia D1 cDNA cloned from kidney and 46% identical to the D1s reported in other vertebrates. Northern blot analysis shows that FhD1 mRNA is expressed in F. heteroclitus liver, consistent with prior biochemical evidence for hepatic D1 activity. Furthermore, heterologous expression of the FhD1 cDNA resulted in a protein with properties similar to the D1-like activity in F. heteroclitus liver. The cloned enzyme, like the native species, is relatively insensitive to inhibition by PTU, but mutation of Ser-159 in FhD1 to the Pro residue found in D2 and D3 isoforms increased the sensitivity to PTU. Our results show that, under basal conditions, killifish liver indeed expresses a D1 enzyme that is homologous to mammalian D1s, establishing this as a useful model in which to study the regulation of D1 and D2 concurrently.
Orozco
Aurea
A
Instituto de Neurobiología, Universidad Nacional Autónoma de México, Campus UNAM-UAQ-Juriquilla, Querétaro 76230, Qro., Mexico. aureao@servidor.unam.mx
Villalobos
Patricia
P
Jeziorski
Michael C
MC
Valverde-R
Carlos
C
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D004251:DNA Transposable Elements / Q000235:genetics; D018076:DNA, Complementary / Q000737:chemistry; D005260:Female; D023781:Fundulidae / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D007453:Iodide Peroxidase / Q000737:chemistry / Q000235:genetics*; D008099:Liver / Q000201:enzymology*; D008297:Male; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009865:Oocytes / Q000201:enzymology; D011441:Propylthiouracil / Q000494:pharmacology; D012333:RNA, Messenger / Q000032:analysis*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D014162:Transfection; D014982:Xenopus laevis
D004251:DNA Transposable Elements; D018076:DNA, Complementary; D012333:RNA, Messenger; D011441:Propylthiouracil; D007453:Iodide Peroxidase