Duplicated zebrafish insulin‐like growth factor binding protein‐5 genes with split functional domains: evidence for evolutionarily conserved IGF binding, nuclear localization, and transactivation activity

Abstract

Insulin-like growth factor binding protein (IGFBP)-5 is a secreted protein that binds to IGF and modulates IGF actions. IGFBP-5 is also found in the nucleus of mammalian cells and has transactivation activity. The structural basis of this transactivation activity and its role in mediating IGF-independent actions are not clear. Here we report that there are 2 igfbp-5 genes in zebrafish and other teleost fish. In zebrafish, igfbp-5a and -5b are expressed in spatially restricted, mostly nonoverlapping domains during early development. The IGF binding site is conserved in both zebrafish IGFBP-5s, and they are both secreted and capable of IGF binding. Both proteins contain a consensus bipartite nuclear localization signal and were found in the nucleus when introduced into cultured cells. Although zebrafish IGFBP-5b possesses transactivation activity, zebrafish IGFBP-5a lacks this activity. Mutational analysis demonstrated that 2 unique amino acids in positions 22 and 56 of IGFBP-5a are responsible for its lack of transactivation activity. These findings suggest that the duplicated zebrafish IGFBP-5s have evolved divergent regulatory mechanisms and distinct biological properties by partitioning of ancestral structural domains and provide new evidence for a conserved role of the IGF binding, nuclear localization, and transactivation domain of this multifunctional IGFBP.

Calcium deficiency-induced and TRP channel-regulated IGF1R-PI3K-Akt signaling regulates abnormal epithelial cell proliferation

“…They were also subjected to ligand blot using DIG-labeled human IGF-1 and IGF-2 as reported previously. 14 Co-immunoprecipitation (IP). Immunoprecipitation was performed using the 4G10 or anti-p85 antibodies following previously published protocol.…”

“…To verify the efficiency and specificity of these MOs for blocking the translation initiation complex on igfbp5a, we constructed a GFP reporter containing partial 5 0 UTR and coding sequence of igfbp5a or igfbp5b. DNA fragments containing 81 bp of 5 0 UTR and 99 bp of coding sequence of igfbp5a, or 91 bp of 5 0 UTR and 99bp of coding sequence of igfbp5b were PCR-amplified and cloned into the pCS2-EGFP 14 at BamHI/ClaI sites. Primers used are shown in Supplementary Table S2.…”

“…14 Its expression pattern resembles that of trpv5/6, which is specifically expressed in NaR cells. 11 IGFs are evolutionarily conserved peptides that have key roles in regulating cell proliferation, growth, differentiation, and survival.…”

Dynamic transcriptional regulation of autocrine/paracrine igfbp1, 2, 3, 4, 5, and 6 in the skeletal muscle of the fine flounder during different nutritional statuses

“…In fish, several studies have described the isolation of individual igfbps (Duan et al 1999, Maures & Duan 2002, Chen et al 2004, Bower et al 2008, Pedroso et al 2009a, Peterson & Waldbieser 2009, Dai et al 2010, Rahman & Thomas 2011.…”

“…Previous studies on the biology of Igfbps in fish have shown that Igfbps regulate embryonic development (Duan et al 1999, Kajimura et al 2005) and exert ligand-dependent and -independent mechanisms (Dai et al 2010, Zhong et al 2011, as well as showing that Igfbps are modulated by Gh treatment (Duan et al 1999, Cheng et al 2002, Chen et al 2004, Pedroso et al 2009b, oxygen availability (Kajimura et al 2005, Rahman & Thomas 2011, Sun et al 2011, reproduction , Chen et al 2010, and nutrition , Bower et al 2008, Pedroso et al 2009a, Peterson & Waldbieser 2009, Amaral & Johnston 2011, Macqueen et al 2011. Nevertheless, studies on local muscle-derived igfbps (autocrine/ paracrine) have been scarce and have recently shown contrasting results, reflecting a complex physiology of Igfbps and suggesting that intricate patterns of regulation have evolved between and within teleost lineages , Bower et al 2008, Amaral & Johnston 2011, Macqueen et al 2011.…”