Abstract
BackgroundFrog metamorphosis is totally dependent on thyroid hormone (T3) and mimics the postembryonic period around birth in mammals. It is an excellent model to study the molecular basis of postembryonic development in vertebrate. We and others have shown that many, if not all, matrix metalloproteinases (MMPs), which cleave proteins of the extracellular matrix as well as other substrates, are induced by T3 and important for metamorphosis. MMP activity can be inhibited by tissue inhibitors of metalloproteinase (TIMPs). There are 4 TIMPs in vertebrates and their roles in postembryonic development are poorly studied.Methodology/Principal FindingsWe analyzed the TIMP2 genes in Xenopus laevis and the highly related species Xenopus tropicalis and discovered that TIMP2 is a single copy gene in Xenopus tropicalis as in mammals but is duplicated in Xenopus laevis. Furthermore, the TIMP2 locus in Xenopus tropicalis genome is different from that in human, suggesting an evolutionary reorganization of the locus. More importantly, we found that the duplicated TIMP2 genes were similarly regulated in the developing limb, remodeling intestine, resorbing tail during metamorphosis. Unexpectedly, like its MMP target genes, the TIMP2 genes were upregulated by T3 during both natural and T3-induced metamorphosis.Conclusions/SignificanceOur results indicate that TIMP2 is highly conserved among vertebrates and that the TIMP2 locus underwent a chromosomal reorganization during evolution. Furthermore, the unexpected upregulation of TIMP2 genes during metamorphosis suggests that proper balance of MMP activity is important for metamorphosis.
Highlights
Matrix metalloproteinases (MMPs) are Zn2+ dependent extracellular or membrane-bound proteinases that can cleave protein components of the extracellular matrix (ECM) as well as nonECM proteins with overlapping substrate specificities [1,2,3,4,5,6,7]
BLAST search with this cDNA sequence identified two additional homologous cDNA sequence entries in X. laevis (GenBank accession #BC074452 and NM_001094279, 99% and 94% identical to AY037944, respectively) and one in X. tropicalis (GenBank accession # NM_001015760, 93% identical in nucleotide sequences to AY037944) (Fig. 1a and Table 2). Both X. laevis clones contain a complete open reading frame (ORF) each encoding proteins of 222 aa or 220 aa, respectively, which are highly homologous to the amino acid sequence of human TIMP2A and B (TIMP2), reflecting duplicated TIMP2 genes in the X. laevis genome
X. tropicalis TIMP2 is highly homologous to both X. laevis TIMP2 genes at either the amino acid and nucleotide sequence levels, sharing over 90% homology (Fig. 1b and Table 2)
Summary
Matrix metalloproteinases (MMPs) are Zn2+ dependent extracellular or membrane-bound proteinases that can cleave protein components of the extracellular matrix (ECM) as well as nonECM proteins with overlapping substrate specificities [1,2,3,4,5,6,7]. They can affect cell fate and behavior by remodeling the microenvironment surrounding the cells and/or altering inter-cellular signaling. We show that both copies of the TIMP2 genes are but unexpectedly upregulated during metamorphosis in different organs that temporally correlates organ specific metamorphic changes. Our findings suggest that proper control of MMP activity is important for temporal regulation of tissue metamorphosis
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