Ribosomal Protein Gene Knockdown Causes Developmental Defects in Zebrafish

The ribosomal proteins (RPs) form the majority of cellular proteins and are mandatory for cellular growth. RP genes have been linked, either directly or indirectly, to various diseases in humans. Mutations in RP genes are also associated with tissue-specific phenotypes, suggesting a possible role in organ development during early embryogenesis. However, it is not yet known how mutations in a particular RP gene result in specific cellular changes, or how RP genes might contribute to human diseases. The development of animal models with defects in RP genes will be essential for studying these questions. In this study, we knocked down 21 RP genes in zebrafish by using morpholino antisense oligos to inhibit their translation. Of these 21, knockdown of 19 RPs resulted in the development of morphants with obvious deformities. Although mutations in RP genes, like other housekeeping genes, would be expected to result in nonspecific developmental defects with widespread phenotypes, we found that knockdown of some RP genes resulted in phenotypes specific to each gene, with varying degrees of abnormality in the brain, body trunk, eyes, and ears at about 25 hours post fertilization. We focused further on the organogenesis of the brain. Each knocked-down gene that affected the morphogenesis of the brain produced a different pattern of abnormality. Among the 7 RP genes whose knockdown produced severe brain phenotypes, 3 human orthologs are located within chromosomal regions that have been linked to brain-associated diseases, suggesting a possible involvement of RP genes in brain or neurological diseases. The RP gene knockdown system developed in this study could be a powerful tool for studying the roles of ribosomes in human diseases.

[1]  The structure and function of eukaryotic ribosomes. , 1979, Annual review of biochemistry.

[2]  D A Kane,et al.  The identification of genes with unique and essential functions in the development of the zebrafish, Danio rerio. , 1996, Development.

[3]  A. Schier,et al.  A genetic screen for mutations affecting embryogenesis in zebrafish. , 1996, Development.

[4]  M. Morishima,et al.  Dominant lethality of the mouse skeletal mutation tail-short (Ts) is determined by the Ts allele from mating partners. , 1998, Genomics.

[5]  A. Lambertsson.  The minute genes in Drosophila and their molecular functions. , 1998, Advances in genetics.

[6]  N Goodman,et al.  A map of 75 human ribosomal protein genes. , 1998, Genome research.

[7]  Peter Gustavsson,et al.  The gene encoding ribosomal protein S19 is mutated in Diamond-Blackfan anaemia , 1999, Nature Genetics.

[8]  R. Stevenson,et al.  X-linked mental retardation with variable stature, head circumference, and testicular volume linked to Xq12-q21. , 1999, American journal of medical genetics.

[9]  I. Dianzani,et al.  Mutations in ribosomal protein S19 gene and diamond blackfan anemia: wide variations in phenotypic expression. , 1999, Blood.

[10]  S. Ekker,et al.  Effective targeted gene ‘knockdown’ in zebrafish , 2000, Nature Genetics.

[11]  A. Tolun,et al.  The novel genetic disorder microhydranencephaly maps to chromosome 16p13.3-12.1. , 2000, American journal of human genetics.

[12]  N. Kenmochi,et al.  A complete map of the human ribosomal protein genes: assignment of 80 genes to the cytogenetic map and implications for human disorders. , 2001, Genomics.

[13]  Rune R. Frants,et al.  Subunits of the translation initiation factor eIF2B are mutant in leukoencephalopathy with vanishing white matter , 2001, Nature Genetics.

[14]  S. Naidu,et al.  Mutations in each of the five subunits of translation initiation factor eIF2B can cause leukoencephalopathy with vanishing white matter , 2002, Annals of neurology.

[15]  Nancy Hopkins,et al.  Insertional mutagenesis in zebrafish rapidly identifies genes essential for early vertebrate development , 2002, Nature Genetics.

[16]  N. Kenmochi,et al.  The human ribosomal protein genes: sequencing and comparative analysis of 73 genes. , 2002, Genome research.

[17]  J. Heasman.  Morpholino oligos: making sense of antisense? , 2002, Developmental biology.

[18]  P. Pandolfi,et al.  Does the ribosome translate cancer? , 2003, Nature Reviews Cancer.

[19]  T. Steitz,et al.  The roles of ribosomal proteins in the structure assembly, and evolution of the large ribosomal subunit. , 2004, Journal of molecular biology.

[20]  S. Karlsson,et al.  Targeted Disruption of the Ribosomal Protein S19 Gene Is Lethal Prior to Implantation , 2004, Molecular and Cellular Biology.

[21]  C. Broeckhoven,et al.  Hot-spot residue in small heat-shock protein 22 causes distal motor neuropathy , 2004, Nature Genetics.

[22]  T. Glaser,et al.  Ribosomal protein L24 defect in Belly spot and tail (Bst), a mouse Minute , 2004, Development.

[23]  Akihiro Nakao,et al.  RPG: the Ribosomal Protein Gene database , 2004, Nucleic Acids Res..

[24]  Nancy Hopkins,et al.  Many Ribosomal Protein Genes Are Cancer Genes in Zebrafish , 2004, PLoS biology.

[25]  Nancy Hopkins,et al.  Identification of 315 genes essential for early zebrafish development. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[26]  K. Xia,et al.  Small heat-shock protein 22 mutated in autosomal dominant Charcot-Marie-Tooth disease type 2L , 2005, Human Genetics.

[27]  Harry F Noller,et al.  RNA Structure: Reading the Ribosome , 2005, Science.

[28]  H. Ropers.  X-linked mental retardation: many genes for a complex disorder. , 2006, Current opinion in genetics & development.

[29]  S. Ellis,et al.  Ribosomes and marrow failure: coincidental association or molecular paradigm? , 2006, Blood.