MECP2 Isoform-Specific Vectors with Regulated Expression for Rett Syndrome Gene Therapy

BACKGROUND:Rett Syndrome (RTT) is an Autism Spectrum Disorder and the leading cause of mental retardation in females. RTT is caused by mutations in the Methyl CpG-Binding Protein-2 (MECP2) gene and has no treatment. Our objective is to develop viral vectors for MECP2 gene transfer into Neural Stem Cells (NSC) and neurons suitable for gene therapy of Rett Syndrome. METHODOLOGY/PRINCIPAL FINDINGS:We generated self-inactivating (SIN) retroviral vectors with the ubiquitous EF1alpha promoter avoiding known silencer elements to escape stem-cell-specific viral silencing. High efficiency NSC infection resulted in long-term EGFP expression in transduced NSC and after differentiation into neurons. Infection with Myc-tagged MECP2-isoform-specific (E1 and E2) vectors directed MeCP2 to heterochromatin of transduced NSC and neurons. In contrast, vectors with an internal mouse Mecp2 promoter (MeP) directed restricted expression only in neurons and glia and not NSC, recapitulating the endogenous expression pattern required to avoid detrimental consequences of MECP2 ectopic expression. In differentiated NSC from adult heterozygous Mecp2(tm1.1Bird)+/- female mice, 48% of neurons expressed endogenous MeCP2 due to random inactivation of the X-linked Mecp2 gene. Retroviral MECP2 transduction with EF1alpha and MeP vectors rescued expression in 95-100% of neurons resulting in increased dendrite branching function in vitro. Insulated MECP2 isoform-specific lentiviral vectors show long-term expression in NSC and their differentiated neuronal progeny, and directly infect dissociated murine cortical neurons with high efficiency. CONCLUSIONS/SIGNIFICANCE:MeP vectors recapitulate the endogenous expression pattern of MeCP2 in neurons and glia. They have utility to study MeCP2 isoform-specific functions in vitro, and are effective gene therapy vectors for rescuing dendritic maturation of neurons in an ex vivo model of RTT

Hafnium-tungsten Chronometry And The Timing Of Terrestrial Core Formation

THE accretion of the Earth and Moon within the solar nebula is thought(1-3) to have taken 50 to 100 million years. But the timing of formation of the Earth's core has been controversial, with some(4,5) proposing that it took place within the first 15 Myr of Earth's accretion history and others(6,7) proposing that it occurred after 50 Myr of accretion. Meteorite chronometry based on the Hf-182-W-182 system has the potential to resolve this debate, as segregation of a metal core from silicates should induce strong fractionation of hafnium from tungsten. Here we report tungsten isotope compositions for two iron meteorites, two carbonaceous chondrites, and a lunar mare basalt. We see clear W-182 deficits in both iron meteorites, in agreement with previous results(4,5). But the data for chondrites are inconsistent with the hypothesis of early core formation, suggesting that both this event and the formation of the Moon must have occurred at least 62+/-10 Myr after the iron meteorites formed.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/62600/1/378771a0.pd