Expression of ribosomopathy genes during Xenopus tropicalis embryogenesis

Abstract

BackgroundBecause ribosomes are ubiquitously required for protein production, it was long assumed that any inherited defect in ribosome manufacture would be embryonically lethal. However, several human congenital diseases have been found to be associated with mutations in ribosome biogenesis factors. Surprisingly, despite the global requirement for ribosomes, these “ribosomopathies” are characterized by distinct and tissue specific phenotypes. The reasons for such tissue proclivity in ribosomopathies remain mysterious but may include differential expression of ribosome biogenesis factors in distinct tissues.MethodsHere we use in situ hybridization of labeled antisense mRNA probes and ultra high temporal resolution RNA-Seq data to examine and compare expression of 13 disease associated ribosome biogenesis factors at six key stages in Xenopus tropicalis development.ResultsRather than being ubiquitously expressed during development, mRNAs of all examined ribosome biogenesis factors were highly enriched in specific tissues, including the cranial neural crest and ventral blood islands. Interestingly, expression of ribosome biogenesis factors demonstrates clear differences in timing, transcript number and tissue localization.ConclusionRibosome biogenesis factor expression is more spatiotemporally regulated during embryonic development than previously expected and correlates closely with many of the common ribosomopathy phenotypes. Our findings provide information on the dynamic use of ribosome production machinery components during development and advance our understanding of their roles in disease.Electronic supplementary materialThe online version of this article (doi:10.1186/s12861-016-0138-5) contains supplementary material, which is available to authorized users.

Paired Box 9 (PAX9), the RNA polymerase II transcription factor, regulates human ribosome biogenesis and craniofacial development

“…In order to model PAX9 based craniofacial development and a role for the neural crest, we depleted pax9 in X . tropicalis embryos, a well-established model for human ribosomopathies [ 70 , 79 , 80 ].…”

“…Additionally, studies in both mice and stage 35 X. laevis embryos support a function for PAX9 in the developing neural crest because pax9 expression during development is highest in the sclerotome and pharyngeal pouches [28,77,78]. In order to model PAX9 based craniofacial development and a role for the neural crest, we depleted pax9 in X. tropicalis embryos, a wellestablished model for human ribosomopathies [70,79,80]. We generated F0 embryos depleted of pax9 by CRISPR-mediated gene modification or injected translation blocking morpholino oligonucleotides (MOs) into X. tropicalis embryos and analyzed their embryonic development compared to uninjected controls.…”

Modulation of the tick gut milieu by a secreted tick protein favors Borrelia burgdorferi colonization

“…The Plasmid was linearized to generate sense and antisense RNA by in vitro transcription using T7 or SP6 polymerase with the Hi Scribe RNA synthesis kit (New England Biolabs) and Digoxigenin-11-UTP (Roche) according to the manufacturer’s instructions. Nymphal ticks fed for 48 h were dissected in MEMFA (1M MOPS, 20 mM EGTA, 10 mM MgSO 4 , 38% Formaldehyde) fixed for one hour, dehydrated in 100% methanol and used to assess presence of bacterial RNA in guts by whole-mount in situ hybridization using Digoxigenin-labeled sense or antisense essentially as described for Xenopus embryos 72 . Hybridized RNA was detected with alkaline phosphatase-conjugated anti-Digoxigenin antibody (Sigma-Aldrich) and the substrate BM purple (Roche).…”

Analysis of cell cycle parameters during the transition from unhindered growth to ribosomal and translational stress conditions

“…Furthermore, the conservation of mechanisms for cell cycle, ribosome biogenesis, and ribosome function suggest that activation of stress-related genes in G1 phase, also could contribute to ribosomopathy diseases such as Blackfan Diamond Anemia. Interestingly, recent results suggest that ribosomopathy genes are also expressed during development [ 86 ], suggesting that mechanisms discovered during extreme ribosome related stress could also contribute to normal development.…”