Zinc finger domains of the Cys-Cys-Cys-His (CCCH) class are evolutionarily conserved proteins that bind nucleic acids and are involved in various biological processes. Nearly 60 CCCH-type zinc finger Show more
Zinc finger domains of the Cys-Cys-Cys-His (CCCH) class are evolutionarily conserved proteins that bind nucleic acids and are involved in various biological processes. Nearly 60 CCCH-type zinc finger proteins have been identified in humans and mice, most have not been functionally characterized. Here, we provide the first in vivo functional characterization of ZC3H4-a novel CCCH-type zinc finger protein. Our results show that although Zc3h4 mutant embryos exhibit normal morphology at E3.5 blastocyst stage, they cannot be recovered at E7.5 early post-gastrulation stage, suggesting implantation failure. Outgrowth assays reveal that mutant blastocysts either fail to hatch from the zona pellucida, or can hatch but do not form a typical inner cell mass colony, the source of embryonic stem cells (ESCs). Although there is no change in levels of reactive oxygen species, Zc3h4 mutants display severe DNA breaks and reduced cell proliferation. Analysis of lineage specification reveals that both epiblast and primitive endoderm lineages are compromised with severe reductions in cell number and/or specification in the mutant blastocysts. In summary, these findings demonstrate the essential role of ZC3H4 during early mammalian embryogenesis. Show less
GABAergic signaling in the CNS is terminated in part through uptake of GABA by GABA transporters. We used the yeast two-hybrid system to identify proteins that associate with the carboxy-terminus of t Show more
GABAergic signaling in the CNS is terminated in part through uptake of GABA by GABA transporters. We used the yeast two-hybrid system to identify proteins that associate with the carboxy-terminus of the neuronal GABA transporter GAT1. We found an interaction between GAT1 and the MAGUK protein Pals1. When coexpressed in COS-7 cells, Pals1 co-immunoprecipitates with GAT1. We demonstrate cellular coexpression of GAT1 and Pals1 in the mouse hippocampus and striatum. Functionally, coexpression of GAT1 and Pals1 in COS-7 cells increases [3H]-GABA uptake by GAT1. The mechanism underlying increased uptake is increased levels of GAT1 protein. We hypothesize that Pals1 contributes to the stability of the GAT1, thus promoting the expression level of the transporter protein. In the CNS, Pals1 may stabilize GAT1 at appropriate levels in specific GABAergic neurons. Show less