Breaking the intergenerational cycle of insecure attachment: a review of the effects of attachment-based interventions on maternal sensitivity and infant security.

In this paper the effectiveness of preventive or therapeutic interventions aiming at enhancing parental sensitivity and children's attachment security is addressed. Sixteen pertinent studies have been reviewed, and 12 studies have been included in a quantitative meta-analysis (N = 869). Results show that interventions are more effective in changing parental insensitivity (d = .58) than in changing children's attachment insecurity (d = .17). Longer, more intensive, and therapeutic interventions appear to be less effective than short-term preventive interventions. Interventions which are effective at the behavioral level may not necessarily lead to changes in insecure mental representations of the parents involved. The implications of changes at the behavioral level (sensitivity; attachment) without accompanying changes at the representational level will be discussed.

One adenosine deaminase allele in a patient with severe combined immunodeficiency contains a point mutation abolishing enzyme activity.

We have cloned and sequenced an adenosine deaminase (ADA) gene from a patient with severe combined immunodeficiency (SCID) caused by inherited ADA deficiency. Two point mutations were found, resulting in amino acid substitutions at positions 80 (Lys to Arg) and 304 (Leu to Arg) of the protein. Hybridization experiments with synthetic oligonucleotide probes showed that the determined mutations are present in both DNA and RNA from the ADA-SCID patient. In addition, wild-type sequences could be detected at the same positions, indicating a compound heterozygosity. Studies with ADA expression clones mutagenized in vitro showed that the mutation at position 304 is responsible for ADA inactivation.

Adenosine deaminase: characterization and expression of a gene with a remarkable promoter.

Cosmid clones containing the gene for human adenosine deaminase (ADA) were isolated. The gene is 32 kb long and split into 12 exons. The exact sizes and boundaries of the exon blocks including the transcription start sites were determined. The sequence upstream from this cap site lacks the TATA and CAAT boxes characteristic for eukaryotic promoters. Nevertheless, we have shown in a functional assay that a stretch of 135 bp immediately preceding the cap site has promoter activity. This 135-bp DNA fragment is extremely rich in G/C residues (82%). It contains three inverted repeats that allow the formation of cruciform structures, a 10-bp and a 16-bp direct repeat and five G/C-rich motifs (GGGCGGG) disposed in a strikingly symmetrical fashion. Some of these structural features were also found in the promoter region of other genes and we discuss their possible function. Knowledge of the exact positions of the intron-exon boundaries allowed us to propose models for abnormal RNA processing that occurs in previously investigated ADA-deficient cell lines.

Introduction of sequences encoding functional human adenosine deaminase into mouse cells using a retroviral shuttle system.

A retroviral packaging system was used to generate a murine virus carrying sequences encoding human adenosine deaminase (ADA). To this end, human ADA cDNA was inserted into the retroviral shuttle vector pZIP-NeoSV(X)1. This vector provides all of the cis-acting sequences necessary for the efficient packaging and transmission of the viral genome as well as a selectable gene for G418 resistance. Transfection of this recombinant plasmid into cells that provide essential virus products (psi-2 cells) yielded cell lines that stably produced virions carrying the coding sequence of human ADA. We have used these virions to infect NIH3T3 cells, which after 48 h synthesized catalytically active human ADA. Furthermore, G418-resistant cell lines were obtained from the virus-infected NIH3T3 cells that stably produced the human ADA enzyme.

Cloning of human adenosine deaminase cDNA and expression in mouse cells.

A previously isolated partial cDNA sequence encoding human adenosine deaminase (ADA) was used to probe a cDNA library prepared from human cultured cell mRNA. Clones containing a combined overlapping length of 1462 bp were isolated and sequenced. One of these was found to include the entire ADA coding region. An open reading frame consisting of 363 codons was identified, predicting a polypeptide of Mr 40762. A mammalian expression plasmid was constructed, positioning the ADA coding sequence to be under transcriptional control of the mouse metallothionein promoter. Transfection of cultured mouse L-cells with this plasmid resulted in the acute expression of human ADA enzymatic activity, as assayed by isoelectric focusing.

Adenosine deaminase (ADA) deficiency in cells derived from humans with severe combined immunodeficiency is due to an aberration of the ADA protein.

In order to determine the molecular basis of adenosine deaminase (ADA) deficiency in cells derived from patients with severe combined immunodeficiency (SCID) disease, we used a human ADA cDNA clone (1) to analyse the organization and transcription of the ADA gene in both normal and ADA-SCID cells. In five lymphoblastoid ADA-SCID cell lines we could detect no deletions or rearrangements in the ADA gene and its flanking sequences. Furthermore, synthesis and processing of ADA mRNA appeared to be normal in the ADA-SCID cells, and ADA-specific mRNA from two ADA-SCID cells could be translated in vitro into a protein with the molecular weight of normal ADA; this protein, however, could hardly be precipitated with an ADA antiserum. The results indicate that in these two ADA-SCID cell lines, the lack of ADA activity is not due to transcriptional or translational defects, but to subtle changes in the configuration of the protein affecting both its enzymatic and immunological characteristics.

Isolation of cDNA clones for human adenosine deaminase.

Clones encoding human adenosine deaminase (ADA) were isolated from a cDNA library made from the lymphoblastoid cell line MOLT-4. The isolation procedure was based on the selection of clones hybridizing with a radioactive probe complementary to an RNA preparation, which had been highly enriched in ADA-specific mRNA. The latter RNA preparation was obtained by size-fractionating MOLT-4 RNA and selecting fractions that were translatable into ADA. The assay for the presence of ADA in the in vitro translation products, was based on immunoprecipitation with a specific anti-ADA serum. The antiserum used was shown to precipitate a 42-kDal protein with the properties of ADA. Positive clones were further screened by means of hybrid-released in vitro translation assays. Two clones were obtained which were able to select mRNA that could be translated into a 42-kDal protein immunoprecipitable with the ADA-antiserum. By use of Southern blots containing DNA from somatic cell hybrids, one of these ADA cDNA clones was assigned to the human chromosome 20 known to contain the ADA gene.