Abstract
North American Indian Childhood Cirrhosis (NAIC) is a rare, autosomal recessive, progressive cholestatic disease of infancy affecting the Cree-Ojibway first Nations of Quebec. All NAIC patients are homozygous for a missense mutation (R565W) in CIRH1A, the human homolog of the yeast nucleolar protein Utp4. Utp4 is part of the t-Utp subcomplex of the small subunit (SSU) processome, a ribonucleoprotein complex required for ribosomal RNA processing and small subunit assembly. NAIC has thus been proposed to be a primary ribosomal disorder (ribosomopathy); however, investigation of the pathophysiologic mechanism of this disease has been hindered by lack of an animal model. Here, using a morpholino oligonucleotide (MO)-based loss-of-function strategy, we have generated a model of NAIC in the zebrafish, Danio rerio. Zebrafish Cirhin shows substantial homology to the human homolog, and cirh1a mRNA is expressed in developing hepatocytes and biliary epithelial cells. Injection of two independent MOs directed against cirh1a at the one-cell stage causes defects in canalicular and biliary morphology in 5 dpf larvae. In addition, 5 dpf Cirhin-deficient larvae have dose-dependent defects in hepatobiliary function, as assayed by the metabolism of an ingested fluorescent lipid reporter. Previous yeast and in vitro studies have shown that defects in ribosome biogenesis cause stabilization and nuclear accumulation of p53, which in turn causes p53-mediated cell cycle arrest and/or apoptosis. Thus, the nucleolus appears to function as a cellular stress sensor in some cell types. In accordance with this hypothesis, transcriptional targets of p53 are upregulated in Cirhin-deficient zebrafish embryos, and defects in biliary function seen in Cirhin-deficient larvae are completely abrogated by mutation of tp53. Our data provide the first in vivo evidence of a role for Cirhin in biliary development, and support the hypothesis that congenital defects affecting ribosome biogenesis can activate a cellular stress response mediated by p53.
Highlights
Infantile cholestasis and/or jaundice results from disorders that disrupt hepatobiliary development, inborn errors of metabolism, toxin exposure and infectious or immune-mediated diseases [1]
Arginine-565 is conserved in the zebrafish homolog (R564), and there are no significant regions in the zebrafish Cirhin protein that are not homologous to CIRHIN
North American Indian Childhood Cirrhosis (NAIC) is an autosomal recessive cholestatic disorder caused by missense mutation of CIRH1A, the human homolog of yeast Utp4
Summary
Infantile cholestasis and/or jaundice results from disorders that disrupt hepatobiliary development, inborn errors of metabolism, toxin exposure and infectious or immune-mediated diseases [1]. The genes affected in these disorders, which collectively have been referred to as cholangiopathies, encode signaling molecules necessary for bile duct development, such as Alagille syndrome [3,4], or proteins necessary for the secretion or modification of bile by hepatocytes or biliary epithelial cells, as seen in progressive familial intrahepatic cholestasis [PFIC 1-3], cystic fibrosis, and arthrogryposis-renal dysfunction-cholestasis syndrome. NAIC presents as neonatal jaundice that resolves spontaneously by age 1 year, but affected individuals have persistent direct hyperbilirubinemia that almost uniformly progresses to portal hypertension and biliary cirrhosis. Liver biopsy at the time of diagnosis typically shows bile duct proliferation with luminal bile plugs and portal fibrosis, findings that are nearly identical to extrahepatic biliary atresia (BA) and consistent with biliary epithelial cell injury. Like patients with BA, most reported NAIC patients develop biliary fibrosis with secondary portal hypertension and liver dysfunction. In a case series reporting 30 patients, 47% had died and 23% had undergone liver transplantation in the first two decades of life; all but one of the remaining living patients had compensated cirrhosis, with the oldest of these patients aged 26 years [6]
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