Knockdown of fbxl10/kdm2bb rescues chd7 morphant phenotype in a zebrafish model of CHARGE syndrome

Abstract

CHARGE syndrome is a sporadic autosomal-dominant genetic disorder characterized by a complex array of birth defects so named for its cardinal features of ocular coloboma, heart defects, choanal atresia, growth retardation, genital abnormalities, and ear abnormalities. Approximately two-thirds of individuals clinically diagnosed with CHARGE syndrome have heterozygous loss-of-function mutations in the gene encoding chromodomain helicase DNA-binding protein 7 (CHD7), an ATP-dependent chromatin remodeler. To examine the role of Chd7 in development, a zebrafish model was generated through morpholino (MO)-mediated targeting of the zebrafish chd7 transcript. High doses of chd7 MO induce lethality early in embryonic development. However, low dose-injected embryos are viable, and by 4 days post-fertilization, morphant fish display multiple defects in organ systems analogous to those affected in humans with CHARGE syndrome. The chd7 morphants show elevated expression of several potent cell-cycle inhibitors including ink4ab (p16/p15), p21 and p27, accompanied by reduced cell proliferation. We also show that Chd7 is required for proper organization of neural crest-derived craniofacial cartilage structures. Strikingly, MO-mediated knockdown of the jumonji domain-containing histone demethylase fbxl10/kdm2bb, a repressor of ribosomal RNA (rRNA) genes, rescues cell proliferation and cartilage defects in chd7 morphant embryos and can lead to complete rescue of the CHARGE syndrome phenotype. These results indicate that CHARGE-like phenotypes in zebrafish can be mitigated through modulation of fbxl10 levels and implicate FBXL10 as a possible therapeutic target in CHARGE syndrome.

Etiology and functional validation of gastrointestinal motility dysfunction in a zebrafish model of CHARGE syndrome

“…By 4 days postfertilization (dpf), the GI tract has developed functionally and structurally into an intestinal bulb, midgut, and hindgut, and demonstrates anterograde and retrograde peristaltic movements, consistent with mature motility patterns . chd7 knockdown using morpholinos recapitulates a CHARGE‐like phenotype in zebrafish larvae, including smaller eyes and defects of the heart, otoliths, developing vertebrae, craniofacial cartilage, and segmental blood vessels . Interestingly, these chd7 morphant larvae also show deformities in the branchiomotor neurons of the hindbrain, as well as decreased projection of the trigeminal (CNV) and facial (VII) nerves .…”

Zebrafish Inner Ear Development and Function

“…Analysis of calcium signaling suggested that Cib2 is required for hair cell specification and function, possibly through the regulation of Ca 2+ in mechanotransduction and calcium homeostasis (Riazuddin et al, 2012). Analysis of the morpholino-injected embryos (morphants) showed otolith abnormalities, semicircular canal defects, and a circling swimming behavior (Patten et al, 2012;Balow et al, 2013). As for Harmonin, the Clarin-1 knockout mouse has no retinal phenotype, and so it is hoped that a zebrafish model for Ush3A will shed light on the role of Clarin-1 in this tissue (Phillips et al, 2013).…”

“…Phillips and colleagues have reported the expression of clarin-1, the causative gene for Ush3A, in sensory cells of the ear, lateral line, and retina of the zebrafish. A general role in control of cell proliferation has been proposed, where the morphants show a reduction in proliferation that can be rescued by inhibiting the rRNA regulator fbxl10 (Balow et al, 2013). CHARGE syndrome (ocular coloboma, heart defects, choanal atresia, growth retardation, genital and ear abnormalities) affects many organ systems and is associated with mutations in the chromodomain-containing, ATP-dependent chromatin remodeller CHD7.…”

Epigenetic Developmental Disorders: CHARGE Syndrome, a Case Study

“…In kismet mutant flies, RNA polymerase II transcriptional elongation is abnormal, suggesting another potential avenue for intervention [33]. Work in zebrafish has also uncovered deficits in ribosomal RNA dysgenesis, and overlapping functions between kismet/CHD7 and other genes that contribute to other craniofacial conditions, including Treacher-Collins syndrome [35,36]. Studies on Xenopus laevis also showed that neural crest derivatives are sensitive to changes in Chd7 dosage and exhibit CHARGE-like features, providing yet another potential route for early developmental intervention [37].…”