The human GCOM1 complex gene interacts with the NMDA receptor and internexin-alpha.

The known functions of the human GCOM1 complex hub gene include transcription elongation and the intercalated disk of cardiac myocytes. However, in all likelihood, the gene's most interesting, and thus far least understood, roles will be found in the central nervous system. To investigate the functions of the GCOM1 gene in the CNS, we have cloned human and rat brain cDNAs encoding novel, 105 kDa GCOM1 combined (Gcom) proteins, designated Gcom15, and identified a new group of GCOM1 interacting genes, termed Gints, from yeast two-hybrid (Y2H) screens.

Determination of a low-level percent enantiomer of a compound with no ultraviolet chromophore using vibrational circular dichroism (VCD): enantiomeric purity by VCD of a compound with three chiral centers.

The chiral configuration of three of the four chiral centers in the investigational drug MLN4924 is locked by an intermediate (1S,2S,4R)-4-amino-2-(hydroxymethyl)cyclopentanol (designated as INT1a). The intermediate INT1a is a key component to the molecule, but its multiple chiral centers and lack of chromophore make it challenging to analyze for chiral purity of the desired enantiomer when it is contaminated with a small amount of its undesired enantiomer. Vibrational circular dichroism (VCD) is a technique that uses the infrared (IR) regions of the electromagnetic spectrum and as INT1a contains IR active groups, we considered using VCD to determine the chiral purity of INT1a.

GRINL1A colocalizes with N-methyl D-aspartate receptor NR1 subunit and reduces N-methyl D-aspartate toxicity.

We hypothesized that proteins from the GRINL1A complex transcription unit called Gcom proteins modulate glutamatergic neurotransmission through interaction with the NR1 subunit of the N-methyl D-aspartate (NMDA) receptor. Cotransfection of hemagglutinin-tagged Gcom1 (GRINL1A combined transcript 1) and NR1 cDNAs into HEK293 cells revealed overlapping fluorescent signals in the plasma membrane. Coimmunoprecipitation studies demonstrated reciprocal coimmunoprecipitation from rat brain protein isolates, suggesting an interaction between GRINL1A proteins and the NMDA receptor.

The human GRINL1A gene defines a complex transcription unit, an unusual form of gene organization in eukaryotes.

Sequencing of genomic DNA and cloned transcripts from the 200-kb human GRINL1A gene on chromosome 15 revealed a complex gene structure comprising at least 28 exons. In one gene model, transcription begins at exon 1 and ends at exon 15b. Another gene model begins transcription at exon 20 and terminates at exon 23, 24, or 28.

Regulation of the glial glutamate transporter GLT-1 by glutamate and delta-opioid receptor stimulation.

The excitatory effect of presynaptically released glutamate is tightly regulated and terminated by high affinity sodium-dependent glutamate transporters. The regulation of the glial glutamate transporter GLT-1 is potentially important in synaptic modulation. Using astroglial cultures prepared from the rat cerebral cortex, we found that the delta-opioid receptor agonist [D-pen2,D-pen5]-enkephalin decreases and glutamate increases the expression of the GLT-1 transporter mRNA.

Gap junctions formed by connexins 26 and 32 alone and in combination are differently affected by applied voltage.

Gap junctions are formed by a family of homologous proteins termed connexins. Their channels are dodecamers, and homomeric forms differ in their properties with respect to control by voltage and other gating stimuli. We report here the properties of coupling from expression of connexin complementary RNAs (cRNAs; sense to mRNA, antisense to cDNA) in Xenopus oocyte pairs in which endogenous coupling was blocked by injection of DNA oligonucleotides antisense to the mRNA of Cx38, the principal endogenous connexin.

Regulated expression of genes inserted at the human chromosomal beta-globin locus by homologous recombination.

We have examined the effect of the site of integration on the expression of cloned genes introduced into cultured erythroid cells. Smithies et al. [Smithies, O.

DNA polymorphisms indicate loss of heterozygosity for chromosome 11 of D98AH2 cells.

Studies with cell hybrids of normal diploid cells fused with tumorigenic D98AH2 (D98) cells had implicated human chromosome 11 of a normal cell as carrying tumorigenicity suppressing information. The cervical carcinoma-derived D98 (HeLa) cells contain two copies of chromosome 11. In this study, analysis of restriction fragment length polymorphism of DNA from D98 cells digested with one of nine restriction endonucleases and hybridized with five DNA probes for highly polymorphic regions on the short arm of chromosome 11 detected no heterozygosity at the insulin (INS), Harvey murine sarcoma virus 1 (HRAS1), and the beta-globin cluster (HBBC) regions.

Coordinate modulation of transfected HSV thymidine kinase and human globin genes.

We have shown that high-frequency phenotypic switching of a transfected gene is associated with alterations in chromatin structure. To examine this phenomenon further, a plasmid containing HSV thymidine kinase and human alpha- and gamma-globin genes was transfected into mouse L cells. All three genes were expressed through utilization of their individual promoters.

Regulated expression of human globin genes and flanking DNA in mouse erythroleukemia--human cell hybrids.

The expression of human globin genes and intergenic DNA is being investigated in MEL-human hybrids containing intact human chromosomes derived from nonerythroid cells. The studies indicate that a factor(s) present in MEL cells can cause activation of both beta- and gamma-globin genes that can be further stimulated by treatment with DMSO, an inducer of MEL cell differentiation. MEL-lymphoblast and MEL-fibroblast hybrids express an adult-like program in which much larger amounts of beta-globin mRNA than gamma-globin mRNA are produced; human beta-globin is at least as efficient as mouse beta-globin gene expression.

Introduction and expression of a fetal human globin gene in mouse fibroblasts.

An 8.5-kilobase segment of cloned human DNA including the complete G gamma-globin gene was introduced into LMTK- cells by the calcium phosphate precipitation method in the presence or absence of carrier DNA. Transfectants containing one or more copies of intact G gamma-globin genes were obtained either by ligation of the human DNA segment to a plasmid containing the herpes simplex virus thymidine kinase gene or by nonligated cotransfer.