Increased Activity of Hypoxia-Inducible Factor 1 Is Associated with Early Embryonic Lethality in Commd1 Null Mice

Abstract

COMMD1 (previously known as MURR1) belongs to a novel family of proteins termed the copper metabolism gene MURR1 domain (COMMD) family. The 10 COMMD family members are well conserved between vertebrates, but the functions of most of the COMMD proteins are unknown. We recently established that COMMD1 is associated with the hepatic copper overload disorder copper toxicosis in Bedlington terriers. Recent in vitro studies indicate that COMMD1 has multiple functions, including sodium transport and NF-B signaling. To elucidate the function of Commd1 in vivo, we generated homozygous Commd1 null (Commd1 ؊/؊ ) mice. Commd1 ؊/؊ embryos died in utero between 9.5 and 10.5 days postcoitum (dpc), their development was generally retarded, and placenta vascularization was absent. Microarray analysis identified transcriptional upregulation of hypoxia-inducible factor 1 (HIF-1) target genes in 9.5-dpc Commd1 ؊/؊ embryos compared to normal embryos, a feature that was associated with increased Hif-1␣ stability. Consistent with these observations, COMMD1 physically associates with HIF-1␣ and inhibits HIF-1␣ stability and HIF-1 transactivation in vitro. Thus, this study identifies COMMD1 as a novel regulator of HIF-1 activity and shows that Commd1 deficiency in mice leads to embryonic lethality associated with dysregulated placenta vascularization.

Distinct Wilson’s Disease Mutations in ATP7B Are Associated With Enhanced Binding to COMMD1 and Reduced Stability of ATP7B

“…Mutations in pEBB-ATP7B(1-650)-Flag and in pEBB-ATP7B-Flag were introduced using the Quickchange site-directed mutagenesis method (Stratagene). Plasmids encoding short hairpin RNAs (shRNAs) were generated by cloning target sequences against COMMD1 (GTCTATTGCGTCTGCAGAC) or ATOX1 (GGTCTGCATTGAATCTGAC) in pRETRO-SUPER as previously described 20 .…”

“…HepG2 cells were transfected using Fugene-6 (Roche, Basle, Switzerland). The generation of stable HEK 293T COMMD1 and ATOX1 knockdown cells has been described elsewhere 20 .…”

“…Previous studies revealed that COMMD1 associates with several proteins to regulate the stability of such interacting proteins 20,21,24 . Therefore, we hypothesized that COMMD1 could also regulate the stability of ATP7B.…”

“…Recent studies have revealed that COMMD1 also functions in a variety of other cellular processes 19 . Analysis of a COMMD1 knockout mouse implicated COMMD1 as a negative regulator of hypoxia-inducible factor 1 (HIF-1) 20 . In addition, inhibitory functions for COMMD1 in sodium transport and NF-κB signaling have been identified 21 -23.…”

Posttranslational regulation of copper transporters

“…These mutations are associated with misfolding, mislocalization of ATP7B to the endoplasmic reticulum, and decreased protein expression due to increased proteasomal degradation. COMMD1 has been implicated in regulation of protein stability of components of the NF-jB and HIF1 signaling pathways [119][120][121][122]. These observations suggest that COMMD1 may affect copper homeostasis by regulating the stability of newly synthesized ATP7B.…”