PMC:4570552 / 17559-19647 JSONTXT

{"project":"2_test","denotations":[{"id":"25865493-11772997-2044025","span":{"begin":269,"end":271},"obj":"11772997"},{"id":"25865493-16470690-2044025","span":{"begin":269,"end":271},"obj":"16470690"},{"id":"25865493-17982426-2044025","span":{"begin":269,"end":271},"obj":"17982426"},{"id":"25865493-19182255-2044025","span":{"begin":269,"end":271},"obj":"19182255"},{"id":"25865493-24664492-2044025","span":{"begin":269,"end":271},"obj":"24664492"},{"id":"25865493-24995868-2044026","span":{"begin":1086,"end":1088},"obj":"24995868"},{"id":"25865493-25561519-2044026","span":{"begin":1086,"end":1088},"obj":"25561519"},{"id":"25865493-24995868-2044027","span":{"begin":1437,"end":1439},"obj":"24995868"},{"id":"25865493-25561519-2044027","span":{"begin":1437,"end":1439},"obj":"25561519"},{"id":"25865493-23656588-2044027","span":{"begin":1437,"end":1439},"obj":"23656588"}],"text":"Because cognitive deficits, with or without brain defects, are invariably present among subjects with MAF mutations affecting the GSK3 recognition motif, but absent in individuals with congenital cataracts caused by MAF mutations involving the DNA binding domain,13,30–33 we posited that expression of the mutant alleles here identified should induce defects in neurogenesis, whereas alleles driving dominant cataracts through a haploinsufficiency model should not. To test this hypothesis in vivo, we analyzed the impact of these mutation classes on the integrity of the central nervous system (CNS) using a zebrafish model. The optic tectum is a major component of the vertebrate midbrain, comprising a structure equivalent to the mammalian superior colliculus, with external layers collecting sensory information and internal layers having a motor-function. We previously showed that measurement of the area of this structure represents a robust surrogate for brain volume and that reduction of the absolute area of the tectum correlates with neurodevelopmental defects in humans.34,35 We injected clutches of 50–100 embryos with human mRNA encoding wild-type MAF, each of four alleles carrying mutations discovered in the present study, or the cataract-associated allele. Experimental work was carried out under protocols approved by the Institutional Animal Care and Use Committee of the Duke University, as previously described.34–36 Following quantitative measurement of the surface area of the tecta (blind to injection cocktail; replicated), we observed that expression of either wild-type and cataract-associated (c.863G\u003eC; p.Arg288Pro) MAF mRNAs did not induce appreciable brain volume differences. In contrast, injection of human MAF mRNA encoding each of the p.Ser54Leu, p.Thr58Ala, p.Pro59Leu, or p.Pro69Arg changes caused a statistically significant reduction of the size of the optic tecta (p \u003c 0.0001; Figure 4). These data were concordant with our in vitro model and provided further evidence for the differential impact of these two mutation classes on CNS development."}